The role of DNA sequence in directing histone core placement on restriction fragments in vitro will be investigated using various histone-DNA assembly procedures. The in vitro assembled nucleosomes will be characterized by nuclease digestion and polyacrylamide gel electrophoresis. The ability of prokaryotic regulatory proteins to bind to nucleosomes containing their regulatory sequences will be investigated with particular emphasis on the role of the nucleosome in accommodating the interaction by changing conformation. Binding interactions will be monitored by sucrose gradient sedimentation of the complexes. The importance of conformational changes will be monitored using cross-linking procedures and nuclease digestion. The interaction of E. coli RNA polymerase with promoter-containing nucleosomes will be studied with respect to the kinetics and mechanisms of binding and the ability to initiate transcription specifically.